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Updated: May 7, 2026

Identification of Kinase-substrate Pairs Using High Throughput Screening

Identification of Kinase-substrate Pairs Using High Throughput Screening

Published on: August 29, 2015

High-throughput screens for eEF-2 kinase.

Ashwini K Devkota¹, Mangalika Warthaka, Ramakrishna Edupuganti

¹1Texas Screening Alliance for Cancer Therapeutics, The University of Texas at Austin, TX, USA.

Journal of Biomolecular Screening

|October 1, 2013

Summary

This summary is machine-generated.

Two new high-throughput assays were developed to screen for inhibitors of eukaryotic elongation factor 2 kinase (eEF-2K), a potential therapeutic target for breast cancer and other diseases. These assays will accelerate the discovery of novel eEF-2K inhibitors.

Keywords:

Sox assay development eEF-2K inhibitors high throughput screen luminescence

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Last Updated: May 7, 2026

Identification of Kinase-substrate Pairs Using High Throughput Screening

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Area of Science:

Biochemistry
Molecular Biology
Pharmacology

Background:

Eukaryotic elongation factor 2 kinase (eEF-2K) is implicated in breast cancer, gliomas, and depression.
The lack of potent eEF-2K inhibitors hinders therapeutic development.
High-throughput assay systems are crucial for identifying small-molecule inhibitors.

Purpose of the Study:

To develop and validate robust, high-throughput assay systems for eEF-2K.
To facilitate the screening of large compound libraries for eEF-2K inhibitors.
To advance the development of targeted therapies for eEF-2K-related diseases.

Main Methods:

Purification of recombinant, tag-free eEF-2K from bacteria.
Development of a fluorescence-based assay using a Sox-based peptide substrate.
Development of a luminescence-based assay measuring residual ATP.
Optimization and miniaturization of assays for a 384-well plate format.

Main Results:

A fluorescence assay demonstrated a 5-fold increase in emission upon substrate phosphorylation.
A luminescence assay showed a signal correlating with remaining ATP.
Both assays were validated in screening formats.
The assays are robust and suitable for large-scale screening.

Conclusions:

Developed universal spectroscopic assays for eEF-2K activity.
These assays enable high-throughput screening for small-molecule inhibitors.
The findings will significantly contribute to developing therapies targeting eEF-2K.